Poly-ubiquitin Binding by the Polyglutamine Disease Protein Ataxin-3 Links Its Normal Function to Protein Surveillance Pathways

Yaohui Chai, Sarah Shoesmith Berke, Robert E. Cohen, Henry L. Paulson

Research output: Contribution to journalArticle

127 Citations (Scopus)

Abstract

In at least nine inherited diseases polyglutamine expansions cause neurodegeneration associated with protein misfolding and the formation of ubiquitin-conjugated aggregates. Although expanded polyglutamine triggers disease, functional properties of host polyglutamine proteins also must influence pathogenesis. Using complementary in vitro and cell-based approaches we establish that the polyglutamine disease protein, ataxin-3, is a poly-ubiquitin-binding protein. In stably transfected neural cell lines, normal and expanded ataxin-3 both co-precipitate with poly-ubiquitinated proteins that accumulate when the proteasome is inhibited. In vitro pull-down assays show that this reflects direct interactions between ataxin-3 and higher order ubiquitin conjugates; ataxin-3 binds K48-linked tetra-ubiquitin but not di-ubiquitin or mono-ubiquitin. Further studies with domain-deleted and site-directed mutants map tetra-ubiquitin binding to ubiquitin interaction motifs situated near the polyglutamine domain. In surface plasmon resonance binding analyses, normal and expanded ataxin-3 display similar submicromolar dissociation constants for tetra-ubiquitin. Binding kinetics, however, are markedly influenced by the surrounding protein context; ataxin-3 that lacks the highly conserved, amino-terminal josephin domain shows significantly faster association and dissociation rates for tetra-ubiquitin binding. Our results establish ataxin-3 as a poly-ubiquitin-binding protein, thereby linking its normal function to protein surveillance pathways already implicated in polyglutamine pathogenesis.

Original languageEnglish (US)
Pages (from-to)3605-3611
Number of pages7
JournalJournal of Biological Chemistry
Volume279
Issue number5
DOIs
StatePublished - Jan 30 2004

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Polyubiquitin
Ubiquitin
Proteins
Carrier Proteins
Ubiquitinated Proteins
polyglutamine
Ataxin-3
Surface Plasmon Resonance
Surface plasmon resonance
Proteasome Endopeptidase Complex
Precipitates
Assays
Cells
Association reactions

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Poly-ubiquitin Binding by the Polyglutamine Disease Protein Ataxin-3 Links Its Normal Function to Protein Surveillance Pathways. / Chai, Yaohui; Berke, Sarah Shoesmith; Cohen, Robert E.; Paulson, Henry L.

In: Journal of Biological Chemistry, Vol. 279, No. 5, 30.01.2004, p. 3605-3611.

Research output: Contribution to journalArticle

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